ProjectSummary/Abstract A key issue in the alcohol field is the lack of knowledge on the discreet neuronal ensembles that are responsible for excessive alcohol drinking in alcohol-dependent subjects. This is a major obstacle for the alcoholfieldbecauseinvestigationsoftheneuronalensemblesthatmediateexcessivealcoholdrinkingwould provide a comprehensive understanding of the neuronal circuits that causally contribute to alcohol dependence.Therecentdevelopmentofpharmacogeneticandoptogenetictoolsthatallowselectivetargeting of specific neuronal ensembles is a tremendous opportunity to bridge this gap in the literature. The central hypothesis of this proposal is that activation of the parabrachial nucleus (PBN) and central nucleus of the amygdala (CeA) during withdrawal is responsible for the recruitment of a set of discreet neuronal ensembles thatarescatteredthroughoutthebrainandultimatelyresponsibleforexcessivedrinkingandtheemergenceof negativeemotionalstatesindependentrats.Weobtainedrobustpreliminaryresultsthatshowthatselectively targetingtheseneuronalensemblesproduceslong-lastingreversalofexcessivealcoholdrinkingindependent rats, identifies a causal relationship between these ensembles, and reveals novel neuronal pathways that contribute to alcohol dependence. Specific Aim 1 characterizes the role of the CeA and PBN withdrawal neuronal ensembles in excessive alcohol drinking in dependent rats. Specific Aim 2 dissects the role of the different CeA CRF pathways in the recruitment of the neuronal ensembles and excessive alcohol drinking in alcoholdependence.Resultsfromthesestudieshavethepotentialtohaveastrongandlastingimpactinthe field because our approach will improve our understanding of the neurobiological mechanisms of alcohol dependenceandidentifynovelneuronalpopulationsandcircuitsthatspecificallycontrolbehaviorsassociated withalcoholdependence.